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PMID:20561257
| Citation |
Yong, Z, Kotur, Z and Glass, AD (2010) Characterization of an intact two-component high-affinity nitrate transporter from Arabidopsis roots. Plant J. 63:739-48 |
|---|---|
| Abstract |
AtNRT2.1, a polypeptide of the Arabidopsis thaliana two-component inducible high-affinity nitrate transport system (IHATS), is located within the plasma membrane. The monomeric form of AtNRT2.1 has been reported to be the most abundant form, and was suggested to be the form that is active in nitrate transport. Here we have used immunological and transient protoplast expression methods to demonstrate that an intact two-component complex of AtNRT2.1 and AtNAR2.1 (AtNRT3.1) is localized in the plasma membrane. A. thaliana mutants lacking AtNAR2.1 have virtually no IHATS capacity and exhibit extremely poor growth on low nitrate as the sole source of nitrogen. Near-normal growth and nitrate transport in the mutant were restored by transformation with myc-tagged AtNAR2.1 cDNA. Membrane fractions from roots of the restored myc-tagged line were solubilized in 1.5% dodecyl-β-maltoside and partially purified in the first dimension by blue native gel electrophoresis. Using anti-NRT2.1 antibodies, an oligomeric polypeptide (approximate molecular mass 150 kDa) was identified, but monomeric AtNRT2.1 was absent. This oligomer was also observed in the wild-type, and was resolved, using SDS-PAGE for the second dimension, into two polypeptides with molecular masses of approximately 48 and 26 kDa, corresponding to AtNRT2.1 and myc-tagged AtNAR2.1, respectively. This result, together with the finding that the oligomer is absent from NRT2.1 or NAR2.1 mutants, suggests that this complex, rather than monomeric AtNRT2.1, is the form that is active in IHATS nitrate transport. The molecular mass of the intact oligomer suggests that the functional unit for high-affinity nitrate influx may be a tetramer consisting of two subunits each of AtNRT2.1 and AtNAR2.1. |
| Links |
PubMed Online version:10.1111/j.1365-313X.2010.04278.x |
| Keywords |
Anion Transport Proteins/genetics; Anion Transport Proteins/metabolism; Arabidopsis/genetics; Arabidopsis/metabolism; Arabidopsis Proteins/genetics; Arabidopsis Proteins/metabolism; Blotting, Western; Cell Membrane/metabolism; Electrophoresis, Gel, Two-Dimensional; Genetic Complementation Test; Luminescent Proteins/genetics; Luminescent Proteins/metabolism; Microscopy, Fluorescence; Multiprotein Complexes/metabolism; Mutation; Nitrates/metabolism; Plant Roots/genetics; Plant Roots/metabolism; Protein Binding; Protoplasts/metabolism |
| edit table |
Significance
Annotations
| Gene product | Qualifier | GO Term | Evidence Code | with/from | Aspect | Extension | Notes | Status |
|---|---|---|---|---|---|---|---|---|
| GO:0015706: nitrate transport |
ECO:0000315: |
P |
Figure 1 shows that ATnar2.1 mutants do not growth on low nitrate in comparison to the wildtype. Additionally growth can be restored to a level that closely parallels WT growth rates by transformation with a myc-tagged AtNAR2.1). Figure 2 shows that mutant lacking the ATNRT2.1 polypeptide exhibit poor growth on low nitrate (also see page 743). |
complete | ||||
|
involved_in |
GO:0015706: nitrate transport |
ECO:0000315: mutant phenotype evidence used in manual assertion |
P |
Seeded From UniProt |
complete | |||
See also
References
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